Immunofluorescence Assay of RAW264.7 Cells

The immunofluorescence (IF) assay was performed on RAW264.7 cells as described previously [12 (link), 48 (link)]. Briefly, the primary antibodies against PCNA (1:50 dilution, Proteintech, USA), Rab7 (1:50 dilution, Proteintech, USA), LAMP1 (1:50 dilution, Proteintech, USA), LC 3 (1:100 dilution, Cell Signaling Technology, USA) and secondary antibodies conjugated with Alexa Fluor 488 and Alexa Fluor 594 dyes (Cat. no. ZF-0513, ZF-0511, and ZF-0512; ZSGB-BIO, China) were used for the test. To observe the superstructures of F-actin, phalloidine conjugated with Alexa Fluor 594 (Sigma, USA) was used to stain the cells for 1 hour at room temperature. The morphology of cell nuclei was shown by DAPI (Sigma, USA) staining for 10 min. The imaging experiments were digitized on laser scanning confocal microscopes (LSM700, Zeiss, Jena, Germany) as described previously [12 (link), 48 (link)]. In addition, the z-stacks plug-in was used to perform the consecutive scans of the cells in the direction of Z-axis.